Transcriptomic and Proteomic Profiling of Rabbit Kidney Cells Infected with Equine Herpesvirus 8

Abstract

The present study investigated the host cell response to EHV-8 infection in rabbit kidney (RK-13) cells through transcriptomic and proteomic approaches. At 24 h post-infection, a total of 2118 differentially expressed genes (DEGs) were identified, with 1338 upregulated and 780 downregulated. At 48 h, 7388 DEGs were detected, with 4342 upregulated and 3046 downregulated genes. Proteomic analysis revealed 932 differentially expressed proteins (DEPs) at 24 h (364 upregulated and 568 downregulated) and 3866 DEPs at 48 h (2285 upregulated and 1581 downregulated). Of these, 237 upregulated and 336 downregulated proteins were common across both time points. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis indicated that the majority of DEGs and DEPs were enriched in key inflammation-related pathways, notably the TNF and NF-κB signaling pathways. Validation of the transcriptomic and proteomic data was performed using RT-PCR and parallel reaction monitoring (PRM), respectively, and confirmed consistent trends for TNFR1, NF-κB p65, and MAP3K8, as reported in the transcriptomic and proteomic screening. These findings suggest that EHV-8 infection may modulate host immune responses by activating the TNF signaling pathway. However, given that RK-13 cells may not fully replicate viral-host interactions in equine species, further in vivo studies in horses and donkeys are required to provide a more comprehensive understanding of the viral pathogenesis in these animals.

Keywords: EHV-8; RK-13 cells; TNF signaling pathway; immune response; proteomics; transcriptomics.

Figure 1

Differential expression analysis of the transcriptome and proteome in the two infection groups. (A) Differentially expressed genes (DEGs) between the two infection groups as compared to the control group. (B) Upregulated and downregulated DEGs shared by the two infection groups. (C) Downregulated DEPs common to both infection groups. (D) Differentially expressed proteins (DEPs) between the two infection groups as compared to the control group. (E) The two infection groups jointly upregulate proteomic DEPs. (F): The two infection groups jointly downregulate proteomic DEPs. Note: R1 represents the control group, R2 represents the 24 h infection group, and R3 represents the 48 h infection group. D1 represents the control group, D2 represents the 24 h infection group, and D3 represents the 48 h infection group. Note: For the comparison of different groups, p-values are those derived from the t-test.

Figure 2

Transcriptome KEGG pathways enrichment analysis. (A) Graph of the common upregulation KEGG enrichment analysis in the transcriptome for the 24 h and 48 h groups. (B) Graph of the common downregulation KEGG enrichment analysis in the transcriptome for the 24 h and 48 h groups. Note: for the comparison of different groups, p-values are those derived from the t-test.

Figure 3

Proteomic KEGG pathways enrichment analysis. (A) Proteomic upregulation KEGG enrichment analysis of the 24 h and 48 h groups together. (B) Proteomic downregulation KEGG enrichment analysis of the 24 h and 48 h groups together. Note: for the comparison of different groups, p-values are those derived from the t-test, and “*” indicates significant differences (p < 0.05), “**” indicates significant differences (p < 0.01), “***” indicates highly significant (p < 0.001).

Figure 4

Joint transcriptome and proteome KEGG pathway enrichment analysis map. (A) Co-upregulation KEGG enrichment analysis plots for 24 h and 48 h groups. (B) Co-downregulation KEGG enrichment analysis in the 24 h and 48 h groups. Note: for the comparison of different groups, p-values are those derived from the t-test, and “*” indicates significant differences (p < 0.05), “**” indicates significant differences (p < 0.01), “***” indicates significant differences (p < 0.001).

Figure 5

RT-PCR results for 48 h infected groups. Note: for the comparison of different groups, p-values are those derived from the t-test, and “*” indicates significant differences (p < 0.05).

Figure 6

Graph of Western blot results of differential proteins in 48 h infection group. (A) WB results. (B) Relative expression map of differential proteins. Note: for the comparison of different groups, p-values are those derived from the t-test, and “*” indicates significant differences (p < 0.05).